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        <title>Primer Design Overview</title>
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        <h1 align="center">Primer Design Overview </h1>
		<p>In addition to a number of quality control capabilities, LRE provides 
		useful insights into the kinetics of PCR amplification that, among other 
		things, have direct relevance to assessing primer pair performance. As 
		such, LRE has been instrumental Developing the design principles that 
		overcome many of the deficiencies currently associate with primer design 
		and testing. Of all the parameters that we have tested, by far the most 
		important impacting primer performance is melting temperature. </p>
		<p>However, 
		before delving into a detailed discussion about primer design, it is 
		essential to first consider the cycling regime of high capacity assays.</p>
		<p>For reasons discussed in the cycle regime section, an early decision 
		was to design primers with a target melting temperature of 70°C in order 
		to allow primer annealing at a temperature high enough to achieve 
		effective enzyme activity. However, as discussed below, the precise 
		primer melting temperature is somewhat arbitrary, because of a large 
		variation in melting temperature produced by different enzyme 
		formulations, which have been to vary over a range of as much as 10°C 
		range. </p>
		<p>The ability to effectively predict melting temperature is further 
		compounded by the proprietary nature of commercial enzyme formulations. 
		Without knowledge of&nbsp; the precise composition of the enzyme 
		formulation, it is impossible to accurately predict the melting 
		temperature of a primer. One approach to circumventing this problem is 
		to take a more arbitrary approach to predicting primer melting 
		temperature. </p>
		<p><b>Primer design programs </b></p>
		<p>Another confounding issue for primer design are the many programs 
		available for primer design, combined with several methods used to 
		calculate primer melting temperature. Indeed, it is not uncommon to find 
		the predicted melting temperature of a primer differs when different 
		primer design programs are used. Although this is certainly do to 
		ineffective prediction for the simplest algorithms, advanced algorithms 
		have been found to generate highly repeatable predictions.&nbsp; </p>
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